IGEM:MIT/2008/Notebook/Yogurt/sequences

Primers

Include sequence, usage, temperatures, concentration, start date.

P1: Forward Primer for Promoter

• • foward sequence: CCGCTTCTAGAGTAATACGACTCACTATAGGGAATACAAGCTACTTGTTCTTTTTGCATACTAGAGATTAAAGAGGAGAAATACTAGATGCAGAAGAAAAAATCCGC
  • length = 110 bp
  • melting temp = 68.8 C
  • GC content % = 37.3

P2: Reverse Primer for Peptide

• • reverse sequence: GTTCTTCTCCTTTACGCATgctGCCCCCGCCgctGCCGCC
  • length = 69 bp
  • melting temp = 71.8 C
  • GC content % = 50.7

TEV Protease cut site

• Amino Acid Sequence : Glu-Asn-Leu-Tyr-Phe-Gln-Gly (Cuts between Gln-Gly)
• Base Pair Sequence: GAAAACCTGTACTTCCAGGGT

P3: Forward Primer for GFP

• • forward sequence: GGCGGCagcGGCGGGGGCagcATGCGTAAAGGAGAAGAAC

• • includes 21 bp of 2nd half of linker, and 19 bp of GFP
  • length = 40 bp
  • melting temp = 66.2 C
  • GC content % = 50%

P4: Reverse Primer for GFP

• • reverse sequence: CTG CAG CGG CCG CTA CTA GTA AGA GAA TAT AAA AAG CCA GAT TAT TAA TCC GGC TTT TTT ATT ATT TTT ATT AGT GGT GAT GGT GAT GAT GTT TGT ATA GTT CAT CCA TGC
  • length = 111 bp
  • melting temp = 69.6 C
  • GC content % = 36.0

P5: Reverse Primer for GFP without Terminator

• • reverse sequence: CTG CAG CGG CCG CTA CTA GTA TTA TTA GTG GTG ATG GTG ATG ATG TTT GTA TAG TTC ATC CAT GC
  • length = 65bp
  • GC content = 44.6 %
  • melting temp = 68.5 ºC

Plasmids

Include sequence, BioBrick #, functional features, digestion map.

PCR Products

Include sequence, template and primers, functional features.

Other

DNA sequence to synthesize - epitope/p1025/linker (6/17/08)

• The signal peptide sequence can be found at the NCBI genome database

• 6/17, updated signal peptide, silent mutations in 1st part of linker (GGC instead of GGT coding for Gly)

sequence: ATGCAGAAGAAAAAATCCGCACGCCATTTGAACAAAGTGGCTGAA TTAGCCGCAGCACTGCTCCTATCAGCGAGTCCACTGGCGGGAACTTTC GACTACAAAGACGACGACGACAAAGGTGGTGGTCAGCTGAAAACCGCT GACCTGCCGGCTGGTCGTGACGAAACCACCTCTTTCGTTCTGGTTGGT GGTGGTTACCCGTACGACGTTCCGGACTACGCTGGCGGCGGCTCTGGC GGCGGCTCTGGCGGCGGCTCTGAAAACCTGTACTTCCAGGGTGGTGGT GGTTCTGGTGGTGGTTCTGGTGGTGGTTCT

Whole Sequence

CCGCTTCTAGAGTAATACGACTCACTATAGGGAATACAAGCTACTT GTTCTTTTTGCATACTAGAGATTAAAGAGGAGAAATACTAGATGCA GAAGAAAAAATCCGCACGCCATTTGAACAAAGTGGCTGAATTAGCC GCAGCACTGCTCCTATCAGCGAGTCCACTGGCGGGAACTTTCCAGT CAGCCGCTTTTGTCCAAGCTGCCAGTCAAGAAACGGACTACAAAGA CGACGACGACAAAGGTGGTGGTCAGCTGAAAACCGCTGACCTGCCG GCTGGTCGTGACGAAACCACCTCTTTCGTTCTGGTTGGTGGTGGTT ACCCGTACGACGTTCCGGACTACGCTGGCGGCGGCTCTGGCGGCGG CTCTGGCGGCGGCTCTGAAAACCTGTACTTCCAGGGTGGTGGTGGT TCTGGTGGAGGCTCTGGTGGAGGCTCTATGCGTAAAGGAGAAGAAC TTTTCACTGGAGTTGTCCCAATTCTTGTTGAATTAGATGGTGATGT TAATGGGCACAAATTTTCTGTCAGTGGAGAGGGTGAAGGTGATGCA ACATACGGAAAACTTACCCTTAAATTTATTTGCACTACTGGAAAAC TACCTGTTCCATGGCCAACACTTGTCACTACTTTCGGTTATGGTGT TCAATGCTTTGCGAGATACCCAGATCATATGAAACAGCATGACTTT TTCAAGAGTGCCATGCCCGAAGGTTATGTACAGGAAAGAACTATAT TTTTCAAAGATGACGGGAACTACAAGACACGTGCTGAAGTCAAGTT TGAAGGTGATACCCTTGTTAATAGAATCGAGTTAAAAGGTATTGAT TTTAAAGAAGATGGAAACATTCTTGGACACAAATTGGAATACAACT ATAACTCACACAATGTATACATCATGGCAGACAAACAAAAGAATGG AATCAAAGTTAACTTCAAAATTAGACACAACATTGAAGATGGAAGC GTTCAACTAGCAGACCATTATCAACAAAATACTCCAATTGGCGATG GCCCTGTCCTTTTACCAGACAACCATTACCTGTCCACACAATCTGC CCTTTCGAAAGATCCCAACGAAAAGAGAGACCACATGGTCCTTCTT GAGTTTGTAACAGCTGCTGGGATTACACATGGCATGGATGAACTAT ACAAACATCATCACCATCACCACTAATAAAAATAATAAAAAAGCCG GATTAATAATCTGGCTTTTTATATTCTCTTACTAGTAGCGGCCGCTGCAG